Neural lipid storage disease (NLSD) is an autosomal recessive disorder characterized by ichthyosis and myopathy. This disorder represents a new type of defect in muscle metabolism and identifying the NLSD defect is likely to elucidate a hitherto unknown, but critical metabolic pathway involving triacylglycerol recycling. In NLSD, triacylglycerol is stored in all types of cells that have been examined. NLSD fibroblasts express the defect and have been well-characterized biochemically. All investigated lipid metabolic pathways are unaltered, including lysosomal and cytosolic lipases, beta-oxidation, and glycerolipid synthesis. These biochemical studies suggest that the cells are either missing an unknown enzymatic function, localizing abnormally a known (lipase) function required for intracellular lipid recycling, or lacking a specific activator or "apoprotein" required for lipid recycling. We propose to "cure" NLSD cells by introducing the normal gene. We will then clone the curative gene. We will construct a cDNA library from mRNA derived from normal cells and enriched for message and encoding the function missing in NLSD. We will transfect NLSD cells with this cDNA, which will be in an expression vector containing an antibiotic resistance gene. We will select transfectants with the antibiotic, and then separate cured cells containing a normal version of the NLSD gene by fluorescence-activated cell sorting (FACS). We will use a fluorescent dye, Nile Red, that will preferentially intercalates into, and fluoresces within, neural lipids of intracellular droplets. We will sort low- fluorescent cells from the NLSD cured cells to amplify curative cDNA inserts by the polymerase chain reaction (PCR). This identity of the putative NLSD cDNA will be confirmed by using it in a pure form to transfect and cure NLSD cells.